The goal of the proposed research is to elucidate the mechanism whereby vasopressin promotes transcellular water and solute movement in responsive epithelial tissues. The specific objectives are a) to determine the role of microtubules and microfilaments in the hydro-osmotic action of vasopressin, b) to explore the role of cytosolic Ca ions and Na-Ca exchange in the action of the hormone, and in the regulation of transepithelial Na transport. A combined functional, biochemical and morphological approach will continue to be employed in studies in the toad bladder in an attempt to determine a) whether microtubules play an active role in the action of vasopressin, b) whether - and, if so, how - microfilaments are involved, c) whether changes in the level of ionized Ca play a role in the action of the hormone on water and Na movement (? regulating microbubule and/or microfilament function, and a rate-limiting step in the Na transport system, respectively, and d) the characteristic and operational role of Na-Ca exchange at the basolateral surface of the epithelial cells. The role of microtubules will be explored in functional and ultrastructural studies on the effects of griseofulvin (an agent that binds to microtubule-associated proteins, and mimics the vasopressin response). Identification and in situ localization of actin and myosin will be pursued using biochemical, histochemical and immunocytochemical methods. Sites of Ca-regulation of vasopressin-induced water movement, and of transepithelial Na transport, will be defined. The effects of procedures believed to raise cytosolic Ca ion activity (quinidine, Ca ionophores, low serosal Na or K, ouabain, La, low pH) on cytoplasmic microtubule content, cellular cAMP levels, and on Na or K, ouabain, La, low pH) on cytoplasmic microtubule content, cellular cAMP levels, and on Na transport parameters, will be examined. The studies will be extended to the mammalian kidney, by exploring the role of cytosolic Ca and Na-Ca exchange and the functional effects of changes in extracellular pH, in the isolated perfused collecting tubule.